INTERACTION OF CAPILLARY ZONE ELECTROPHORESIS WITH A SHEATH FLOW CUVETTE DETECTOR
INTERACTION OF CAPILLARY ZONE ELECTROPHORESIS WITH A SHEATH FLOW CUVETTE DETECTOR
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DOI：
10.1021/ac00204a014
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发表时间：
1990-03-01
影响因子：
7.4
通讯作者：
DOVICHI, NJ
中科院分区：
文献类型：
--
作者：
CHENG, YF;WU, SL;DOVICHI, NJ
在带鞘流检测器的毛细管区带电泳系统中，加压检测器引起与电泳相反的流动分量。这种压力诱导流动由于其速度分布的不均匀性，会使分离恶化。然而，理论预测和实验验证，在正常操作条件下，检测器不会降低分离。在pH 9和10下的谱带增宽的主要来源是由于纵向扩散，尽管进样体积可限制系统的性能。当进样量最小时，在10-mln分离中，可获得超过250万个理论塔板用于分析氨基酸的硫代氨基甲酰荧光素衍生物。此外，可以以良好的精度测量扩散系数和电泳迁移率。毛细管区带电泳的低流速影响鞘流池的性能。理论预测和实验验证了鞘流检测器毛细管区带电泳峰高随分离电压线性增加。对于径向扩散的分析物，本文还提出了鞘流比色皿中样品流半径的模型。毛细管区带电泳是一种特别有效的离子分析技术（1-10）。在该技术中，分离是基于在电场的影响下，分析物在填充缓冲液的毛细管中的差异迁移。因为分离过程中产生的理论塔板数与所加电压成正比，所以很高的电位可以产生很有效的分离。在该实验室中，理论塔板数大于106，分离电位为30 kV。毛细管区带电泳的成功应用需要仔细注意尽量减少谱带展宽的来源。为了达到尽可能高的分离效率，必须使进样体积、分析物浓度和检测体积最小化。为了获得最佳的分离性能，必须注入非常少量的分析物（通常小于飞摩尔）（1）。鞘流比色皿非常适合小体积、低浓度样品的激光诱导荧光检测（11-17）。尽管比色皿最常见于为流式细胞术的生物医学技术设计的仪器中（18），但该装置的分析应用包括液相色谱法中的激光诱导荧光检测（11）、流动注射分析中的激光诱导荧光检测（12）、纯溶液的研究中的激光诱导荧光检测（13-15）和毛细管区带电泳中的激光诱导荧光检测（16，17）和纯溶液的折射率检测（19，20），毛细管区带电泳中的折射率梯度检测（21），以及纯溶液的热光吸光度检测（22）。类似的装置已用于毛细管区带电泳中的柱后标记（23，24）。
In a capillary zone electrophoresis system with a sheath flow detector, the pressurized detector Induces a component of flow opposite to that of electrophoresis. This pressure-induced flow, because of Its nonuniform velocity profile, can degrade the separation. However, theory predicts and ex-periment verifies that, under normal operating conditions, the detector does not degrade the separation. The major source of band broadening at pH 9 and 10 Is due to longitudinal diffusion, although Injection volume can limit the performance of the system. When Injection volume Is minimized, over 2.5 million theoretical plates are obtained for the analysis of fluorescein thiocarbamyl derivatives of amino acids In a 10-mln separation. Also, diffusion coefficients and electrophoretic mobilities may be measured with good accuracy. The low flow rate of capillary zone electrophoresis affects the performance of the sheath flow cuvette. Theory predicts and experiment verifies that the peak height In capillary zone electrophoresis with the sheath flow detector Increases line-arly with separation voltage. A model Is also presented for the sample stream radius expected In the sheath flow cuvette for analytes that undergo radial diffusion.Capillary zone electrophoresis is a particularly powerful technique for the analysis of ions (1-10). In the technique, separation is based on differential migration of analyte in a buffer-filled capillaryunder the influence of an electric field. Because the number of theoretical plates produced during the separation is proportional to the applied voltage, veryefficient separations may be produced by very high potentials. In this laboratory, theoretical plate counts greater than 106 are rou-tinely produced with separation potentials of 30 kV. Successful application of capillary zone electrophoresis requires careful attention to minimize sources of band broadening. To achieve the highest possible separation ef-ficiency, it is necessary tominimize the injection volume, analyte concentration, and detection volume. Very small amounts of analyte, often less than a femtomole, must be injected to obtain optimum separation performance (1). The sheath flow cuvette is well suited for laser-induced fluorescence detection of small-volume, low-concentration samples (11-17). Although the cuvette is most commonly found in instrumentation designed for the biomedical technique of flow cytometry (18), analytical applications of the device have included laser-induced fluorescence detection in liquid chromatography (11), in flow injection analysis (12), in the study of neat solutions (13-15), and in capillary zone electrophoresis (16, 17) and refractive index detection of neat solutions (19, 20), refractive index gradient detection in ca-pillary zone electrophoresis (21), and thermooptical absorbance detection of neat solutions (22). A similar device has been used for postcolumn labeling in capillary zone electrophoresis (23, 24).